The Ecotoxicological Impacts of Carbamazepine: Environmental Contamination vs. Effects on Freshwater Aquatic Organisms

Abstract

The global population is expected to surpass nine billion by 2050, resulting in increased resource consumption and environmental degradation. This raises concerns about environmental contamination, a topic highlighted since Rachel Carson’s Silent Spring (1962), which warned about Contaminants of Emerging Concern (CECs), including pharmaceuticals. Carbamazepine (CBZ) is a common pharmaceutical contaminant in freshwater environments. In this context, this work presents a review of the effects of this substance on freshwater aquatic organisms-including Danio rerio, Cyprinus carpio, Pimephales promelas, Jenynsia multidentata, Gobiocypris rarus, Tinca tinca, Daphnia magna, Daphnia similis, Corbicula fluminea, Dreissena polymorpha, Pseudokirchneriella subcapitata, Schmidtea mediterranea, and Eriocheir sinensis-based on the analysis of 26 scientific articles. The studied species exhibited various impacts, with Danio rerio showing notable changes in embryo hatching time. Fish studies typically involved CBZ concentrations above 100 µg/L, while invertebrates were exposed to lower levels. Common toxic effects included morphological alterations, behavioral changes, and oxidative stress. To reduce these impacts, advanced wastewater treatment technologies capable of fully removing pharmaceuticals are needed. Pharmacists also play a key role in promoting rational drug use and proper disposal to ensure environmentally safe elimination. Overall, raising awareness and adopting sustainable practices are essential to protect aquatic ecosystems and biodiversity.

Keywords:
Emerging contaminants; Carbamazepine; Ecotoxicology.

INTRODUCTION

The increasing demand for food production and energy generation, driven by population growth and current consumer market demands, has created the need to develop large-scale industrial products that meet the standards of modern society. According to the latest United Nations (UN) report published in 2019, it is estimated that the global population will exceed nine billion people by 2050. This exponential population growth will certainly directly influence excessive resource consumption, increased waste emissions, and higher rates of air, soil, and water pollution, generating challenges related to the release of synthetic chemicals into the environment (UN, 2019).

According to EUROSTAT (Statistical Office of the European Union), in 2022, 200 million tons of chemicals were produced in the 28 member states of the European Union (EU), of which 190 million tons were considered hazardous to health. The production of these compounds is an indicator of potential negative impacts on human health and the environment and should be used as a basis for environmental risk assessment.

Exposure to these chemicals can induce a variety of adverse effects on exposed organisms, including humans, such as endocrine disruption, hepatotoxicity, neurotoxicity, cardiorespiratory effects, among others. In recent years, various studies have investigated these effects, highlighting the relationship between environmental factors and their consequences for exposed organisms. Various chemicals, including bisphenol A (BPA), phthalates, dyes, pesticides, pharmaceuticals, and others, have been shown to alter neurobehavioral health and human reproduction (Sharma et al., 2020). In most cases, contact with these compounds can occur through wastewater, air, or bioaccumulated elements in consumed foods, mainly of animal origin (Encarnação et al., 2019).

Thus,environmentalcontaminationwithcompounds potentially hazardous to human and environmental health has been a concern of the international scientific community for years. In 1962, Rachel Carson published the book "Silent Spring", which is considered a milestone for Environmental Toxicology, because, since that time, environmental movements were initiated. In her book, Rachel Carson highlighted the impact of the indiscriminate release of chemical substances into the environment, focusing primarily on the effects of the pesticide dichlorodiphenyltrichloroethane (DDT) on organisms, especially birds (Carson, 1962; Oliveira, Kummrow, 2021). This book was later widely recognized by regulatory agencies, culminating in the restriction of DDT use during the Stockholm Convention, which was organized to discuss the risks of persistent organic pollutants (Gobbo, 2016). This event was one of the first to address the severity of so-called “emerging contaminants” (EC), substances that are not monitored but are found in the environment and potentially harmful to humans and/or wildlife (Montagner, 2018). These pollutants are currently better known as “contaminants of emerging concern” (CEC), as these compounds are not necessarily new and may have been present in the environment for a considerable time, but concern regarding their presence has only arisen recently (Sauvé, Desrosiers, 2014). It is also worth noting that the terms EC or CEC are not related solely to substances of anthropogenic origin but also include natural compounds with previously unrecognized adverse effects on ecosystems, such as algal toxins (Dorta, Oliveira, 2021; USEPA, 2008).

Currently, the list of these contaminants includes pharmaceuticals and hormones, pesticides, artificial sweeteners, personal care products, disinfection byproducts, perfluorinated compounds, illicit drugs, ultraviolet (UV) filters, flame retardants, among others. In addition to their structural diversity, another important characteristic of these compounds is that many are present in the environment at ng/L and µg/L concentrations, and, therefore, are also known as micropollutants (Dorta, Oliveira, 2021; Schwarzenbach et al., 2006; USEPA, 2008). One of the biggest issues concerning this topic is the lack of regulations regarding the determination of permissible maximum concentrations of these compounds, resulting in few monitoring or control actions to prevent the disposal of these pollutants (Quesada et al., 2019). According to Quesada and collaborators (2019), CECs are generally bioaccumulative and active, and the rate of release of these compounds into the environment tends to increase as the population grows, making the discussion on this subject even more necessary.

In the aquatic environment, CECs are introduced through various means, such as the direct discharge of wastewater, leaching from landfills, runoff from agricultural and urban areas, hospital waste, irregular disposal of garbage and consumer goods, and sewer overflow, potentially reaching drinking water due to the stability of contaminants, which may not be completely eliminated by conventional methods in water treatment plants (Quesada et al., 2019; Kumar et al., 2022; Tran, Reinhard, Gin, 2018; Wieczorko Barán, De Aquino, Lima Sanson, 2023).

Among the mentioned contaminants, pharmaceuticals stand out, as they are administered, absorbed, metabolized, and finally excreted in their original, associated forms or as metabolites (Souza, Aquino, Silva, 2020). Thus, they reach the aquatic environment mainly through the discharge of domestic wastewater, with significant contamination also from industrial and hospital effluents. Additionally, it is important to consider the improper disposal of expired or unused medications in common trash or by flushing them down toilets. This contamination occurs because conventional treatments commonly used in sewage treatment plants cannot completely remove these compounds (Almeida et al., 2021; Bisognin, Wolff, Carissimi, 2018; Tejada, Quiñonez, Peña, 2014).

The first reports of the presence of pharmaceuticals in aquatic environments date back to the 1970s-1990s, when caffeine was detected in surface and wastewater in the United States and the discovery of tetracycline and theophylline in rivers in the United Kingdom. Among the classes of pharmaceuticals present as contaminants, antibiotics and, subsequently, analgesics prevail, but the data vary according to the region and country (Aus der Beek et al., 2016; Patel et al., 2019). However, toxicity assessment studies indicate that the most impactful classes for the aquatic ecosystem are antidepressants, antibiotics, antipsychotics, cardiovascular medications, antineoplastics, and hormones (natural or synthetic). The presence of these pharmaceutical residues in water bodies can cause damage to aquatic ecosystems (such as acute and chronic toxicity, genotoxicity, selection of multi-resistant bacteria, feminization of male fish, and reproductive alterations). Additionally, they are also harmful to human health, and exposure to low doses through water consumption has been related to increased cancer incidence, reduced sperm count, infertility, spontaneous abortions, and risks associated with bacterial resistance to antibiotics (Bisognin, Wolff, Carissimi, 2018).

Among the pharmaceuticals with high polluting potential, this work will address one of the most relevant in aquatic contamination: Carbamazepine (CBZ) (Figure 1), an antiepileptic drug primarily used as an anticonvulsant, but also as a mood stabilizer (Mezzelani, Gorbi, Regoli, 2018). Epilepsy is characterized by a temporary alteration in brain function, generated naturally. For some time, a part of the brain emits incorrect signals, which may be restricted to that location (classified as partial seizure) or involve both hemispheres of the brain (classified as generalized seizure) (Ministério da Saúde do Brasil, 2022).

FIGURE 1
Structure of Carbamazepine (CBZ).

Ambrósio and collaborators (2002) presented various data on CBZ, including its structure, derived from iminodibenzyl (similar to that of tricyclic antidepressants) and its effectiveness, which varies between approximately 30% and 40% in patients with epilepsy. Furthermore, they describe its target as voltage-dependent sodium channels, reducing the frequency of repetitive and sustained action potentials in neurons and, consequently, depressing excessive electrical activity in the brain. Considering its mechanism of action, CBZ demonstrates efficacy in tonic-clonic seizures, being a first-choice drug for these cases, but it can also intensify seizures or induce status epilepticus in other subsyndromes (Betting, Guerreiro, 2008).

Additionally, according to Bertilsson's study (1978), CBZ has an oral bioavailability of over 70% and a high rate of binding to plasma proteins during its distribution (between 70% and 80%). Its metabolism occurs predominantly in the liver, generating metabolites such as carbamazepine-10,11-epoxide, which are excreted in the urine. In this context, approximately 2% of the drug is excreted unchanged.

In the review published by Mezzelani, Gorbi, and Regoli (2018), CBZ was the psychiatric drug with the highest maximum concentration in aquatic environments among others belonging to the same group. This is highly concerning considering that conventional water treatment methods eliminate only about 7% of this compound (Hu et al., 2009). Moreover, it is important to highlight that epilepsy affects 50 million people worldwide and 2% of the Brazilian population (Ministério da Saúde do Brasil, 2022). These high numbers of patients indicate that if they use CBZ, they may unintentionally cause environmental pollution due to the possibility of improper disposal or even during the use of the drug through renal excretion.

In this context, this work aimed to review the impacts of carbamazepine release on aquatic ecosystems, with an emphasis on the effects on the health of freshwater organisms. Additionally, it seeks to raise awareness among pharmaceutical professionals and patients about the importance of proper medication disposal to minimize the environmental impacts associated with drug use and, indirectly, to promote the rational use of medications.

METHODOLOGY

Using the PubMed database, the initial search was conducted using the terms “Carbamazepine contamination” and results published between 2019 and 2024 were selected, generating a total of 475 articles.

Subsequently, a new search was conducted using the terms “Carbamazepine aquatic environment fish”, selecting results between 2016 and 2024, extending the time period as it was a more restricted search. Of the 78 articles found, 30 duplicates were eliminated, generating 48 new results. Finally, the results found on PubMed totaled 523 articles.

To select only the articles that fit this work, 6 keywords were chosen to perform a new filter, optimizing the selection of the final articles: “effects”, “aquatics”, “marine”, “fish”, “environments” and “freshwater”. This method, combined with individual selection from reading the abstracts of each, resulted in a final number of 17 articles found.

To increase the number of results, the “Research Rabbit” site was used to find articles similar to the expected results for further analysis. For this, the work of Calcagno and collaborators (2016) was selected as a standard so that new works with a similar approach and similar authors would be indicated by the site. This yielded a total of 1644 results. Manually filtering those from the period between 2016 and 2024, 80 works were separated and, after individually analyzing each one, 18 more articles were selected for further analysis.

Finally, with a total of 35 selected works, the direction of the work was defined to discuss the effects of CBZ on freshwater organisms, present in more than 71% of the total selected works for this review.

RESULTS

A total of 26 articles were included in this review, representing the complete set of publications retrieved through the search strategy described in the Methodology section. The selected studies provided data on the toxicity of carbamazepine in various freshwater species, including Danio rerio, Cyprinus carpio, Pimephales promelas, Jenynsia multidentata, Gobiocypris rarus, Tinca tinca, Daphnia magna, Daphnia similis, Corbicula fluminea, Dreissena polymorpha, Pseudokirchneriella subcapitata, Schmidtea mediterranea, and Eriocheir sinensis-totaling 13 different organisms. To facilitate a clearer and more organized discussion, the results are presented below, in Figure 2, by species.

FIGURE 2
Number of articles that address each species in this study. Source: Author.

Danio rerio

Zebrafish (Danio rerio) was the most frequently studied freshwater species regarding the effects of CBZ exposure. The reviewed studies addressed multiple endpoints, including development, gene expression, behavior, neurotoxicity, reproductive capacity, and oxidative stress. While some effects were consistently reported, others varied significantly, suggesting that methodological and experimental differences - such as compound concentration, exposure duration, developmental stage, and mixture with co-contaminants - may underlie these discrepancies.

Developmental effects, particularly on embryo hatching, showed contrasting results. Rodrigues et al. (2023) observed that only the metabolite 10,11-dihydrocarbamazepine (diCBZ) reduced the hatching range by 41% at 72 hours post-fertilization (hpf) at 1 µg/L. In contrast, the parent compound CBZ and other metabolites did not significantly affect hatching under the tested conditions. Similarly, most other studies (Chen et al., 2020; Zhou et al., 2019) reported no significant effect or only minor changes. However, Qiang et al. (2016) found that CBZ exposure shortened the hatching time. Such variation may reflect differences in exposure windows, compound formulations (pure CBZ vs. metabolites), or endpoints used to define hatching success.

Gene expression analysis revealed consistent interference by CBZ in several key physiological processes. Rodrigues et al. (2023) documented alterations in genes related to organogenesis and neurotransmission (e.g., raraa, sert, mao, pparg, drd2b), while Qiang et al. (2016) reported increased expression of neurodevelopmental markers (gfap, huC, neuroD, ngn1) at concentrations between 1-5 μg/L. Chen et al. (2020) focused on neurotransmitter pathways, observing inhibition in most genes related to GABA and glutamate systems, including gabra1, grin1b, gria2b, gad1b, and abat, with concentration-dependent effects. Notably, some genes, such as gad1b, showed variable responses depending on the dose (e.g., inhibition at both low and high doses, but not intermediate), suggesting non-monotonic effects.

Yang et al. (2023) explored the impact of CBZ on genes related to antibiotic resistance and intestinal immunity. At low concentrations (0.3 g/ kg), CBZ induced overexpression of resistance genes (aminoglycosides, β-lactams, MDR), while higher concentrations (3 g/kg) led to decreased expression.

After 28 days, CBZ suppressed TLR2 and TJP2a expression - genes involved in mucosal immunity and barrier integrity - and increased pro-inflammatory markers IL-6 and TNFα, suggesting intestinal damage and immunological stress. Together, these studies point to broad transcriptomic effects of CBZ across diverse biological systems, even at environmentally relevant concentrations.

Behavioral and physiological changes were also frequently reported but varied depending on dose and timing. Rodrigues et al. (2023) showed that CBZ and its epoxide metabolite (CBZep) increased the frequency of malformations, particularly in the yolk sac, and impaired circulation, tail detachment, and stimulus response. Qiang et al. (2016) observed accelerated development, premature swim bladder inflation, body elongation, and increased sensitivity to stimuli. However, Chen et al. (2020), even using higher concentrations (up to 100 μg/L), reported no significant structural changes, though they noted reduced spontaneous embryonic movement at 10 μg/L. Zhou et al. (2019) observed increased larval swimming speed, whereas Weichert et al. (2017) found reduced embryonic activity and increased edema frequency. These differences may reflect species strain, developmental stage, or subtle experimental design differences.

Reproductive parameters also yielded divergent findings. Fraz et al. (2018) showed substantial reductions in fecundity (44-66%) in two separate experiments, while Da Silva Santos et al. (2018), using similar concentrations (10 μg/L), found no significant effect on egg production. However, the latter did report reduced viability and increased food intake time on specific days (21, 42, and 63), indicating subtle effects on reproductive efficiency and behavior.

Neurochemical and enzymatic effects were studied by multiple groups. Chen et al. (2020) reported increased AChE activity at 10 and 100 μg/L and GABA enhancement only at the lowest dose (1 μg/L). Glutamate activity was reduced at 1 and 100 μg/L but not at 10 μg/L, again highlighting non-linear dose-response relationships. Jia et al. (2020), however, found that AChE was inhibited at ≥100 μg/L, especially when CBZ was combined with copper (Cu), suggesting possible interaction effects between pharmaceuticals and heavy metals in aquatic environments.

Oxidative stress markers, such as SOD, CAT, and IHR, were commonly investigated. Jia et al. (2020) showed that isolated CBZ at high doses reduced SOD activity, but its co-exposure with Cu increased SOD at moderate concentrations and suppressed it at high doses. CAT was inhibited by CBZ alone and in combination with Cu, particularly at higher concentrations. Da Silva Santos et al. (2018) confirmed CAT inhibition in liver and gills and AChE elevation in both head and muscle tissues, aligning with Chen et al. (2020) but contrasting with Jia et al. (2020). The tissue-specific responses underscore the complexity of interpreting enzymatic endpoints, especially under realistic mixture scenarios.

In summary, despite variations across individual endpoints, there is consistent evidence that CBZ - even at low concentrations - can alter gene expression, impair neurodevelopment, and induce physiological and behavioral changes in Danio rerio. Divergences between studies may be attributed to differences in exposure conditions, developmental stages analyzed, type of CBZ compound (parent vs. metabolites), presence of co-contaminants, and analytical sensitivity. These findings emphasize the need for harmonized protocols and more comparative studies to clarify CBZ’s ecotoxicological mechanisms in aquatic vertebrates.

Cyprinus carpio

Gheorghe et al. (2016) investigated the effects of CBZ on Cyprinus carpio, reporting an LC₅₀ ^of approximately 42.6 mg/L, making it the most lethal compound among those tested, which included diclofenac, acetaminophen, and caffeine.

Gasca-Pérez et al. (2019) evaluated CBZ bioconcentration in different body tissues and found the highest accumulation in the gills (63.42 μg/kg), followed by the brain (42.25 μg/kg) and liver (41.44 μg/kg), after exposure to approximately 147.42 μg/g of the drug in water. The authors also assessed oxidative stress biomarkers at different time points (12 to 96 h), observing increased lipid peroxidation in the liver (70.12% at 12 h and 78.90% at 24 h) and in the gills (75.25% at 96 h). In contrast, a significant reduction was detected in the brain at 24 h (32.45%), 48 h (44.50%), and 72 h (39.86%).

Regarding hydroperoxide content, all tissues exhibited marked increases: in the liver (e.g., 483.65% at 48 h), gills (e.g., 571.14% at 12 h), and brain (e.g., 749.55% at 12 h). The activity of superoxide dismutase (SOD) decreased in all tissues, particularly in the liver (up to 73.84% inhibition at 48 h) and gills (up to 88.84% at 96 h). Similarly, catalase (CAT) activity declined in the liver (e.g., 91.02% at 12 h), gills (e.g., 56.27% at 96 h), and especially in the brain (up to 96.17% at 12 h). These results suggest that CBZ induces strong, time-dependent oxidative stress in C. carpio, with varying degrees of tissue sensitivity.

Pimephales promelas

Zind et al. (2021) assessed the toxicity of CBZ and its metabolites/degradation products in Pimephales promelas. Among the compounds tested, dibenzazepine showed the highest toxicity (LC₅₀ = 3.27 mg/L), followed by CBZ alone (LC₅₀ = 37.30 mg/L). The mixture of all components resulted in an LC₅₀of 41.50 mg/L-lower than CBZ alone but still 3 to 5 times more toxic than the other metabolites tested individually. These findings highlight that not only CBZ but also its transformation products pose relevant ecotoxicological risks.

Jenynsia multidentata

Calcagno et al. (2016) explored the behavioral effects of CBZ in Jenynsia multidentata exposed to different concentrations (10, 50, and 200 μg/L) under stress conditions induced by lighting. An increase in the time spent in the light-exposed compartment was observed in all stress scenarios, including CBZ-stress combinations, which deviates from the species' normal behavior.

Average swimming speed increased under isolated stress but decreased at higher CBZ concentrations, particularly at 200 μg/L. The defense behavior known as freezing showed an increasing trend under stress, except at the highest CBZ concentration, where it was reduced. Time spent at the bottom of the tank was highest in the stress-only group and the 50 μg/L + stress group. In the 10 μg/L + stress group, behavior resembled that of the control, while the 200 μg/L group showed an intermediate response.

The authors suggest that lower CBZ concentrations may reduce stress-related behavioral responses, possibly through modulation of cortisol metabolism. The study raises the hypothesis that CBZ may alter the biotransformation of this stress hormone, thereby influencing the organism’s behavioral coping mechanisms.

Gobiocypris rarus

Yan et al. (2018) exposed Gobiocypris rarus to CBZ concentrations of 1, 10, and 100 μg/L over 28 days and evaluated gene expression alterations. A total of 146 genes showed significant changes in mRNA expression in both sexes. In the liver, 111 genes were altered in females and 71 in males. Further analysis revealed that 36 genes were commonly affected in both sexes at the highest concentration (100 μg/L), with 21 upregulated and 12 downregulated. The affected genes were primarily associated with the hypothalamic-pituitary-gonadal (HPG) axis, lipid metabolism, calcium signaling pathways, among others.

The study also assessed two key steroid hormones, 17β-estradiol (E2) and 11-ketotestosterone (11-KT), as well as vitellogenin (VTG), a precursor protein essential for egg production (Girish, Swetha, Reddy, 2014). Plasma levels of 11-KT were suppressed in both males and females at all concentrations, while E2 levels were altered only in females exposed to 10 and 100 μg/L. VTG levels increased across all treatments, except in males exposed to 1 μg/L of CBZ. These findings suggest potential endocrine-disrupting effects of CBZ even at environmentally relevant concentrations.

Tinca tinca

Stancova et al. (2017) investigated the effects of CBZ alone and in combination with ibuprofen and diclofenac on oxidative metabolism in early life stages of Tinca tinca. CBZ alone (at 60 µg/L) stimulated only one antioxidant enzyme-glutathione peroxidase. However, the drug mixture (administered at 0.02, 0.2, 2, 20, and 60 µg/L in a 1:1:1 ratio) did not induce significant oxidative responses, suggesting no combined pro-oxidant effect during early development. These results indicate that while CBZ alone may modulate specific antioxidant defenses, its interaction with other common pharmaceuticals may not produce additive or synergistic oxidative damage at early life stages.

Daphnia magna

Gheorghe et al. (2016) reported that CBZ had the second-lowest EC₅₀ value (21.87 mg/L) among the substances tested on Daphnia magna, indicating high toxicity. However, Oropesa, Floro, and Palma (2016) found a higher EC₅₀ value (58.54 mg/L), highlighting variability across studies. Complementarily, Zind et al. (2021) determined an LC₅₀ of 111.00 mg/L for this species. Notably, among CBZ metabolites, dibenzazepine showed the highest toxicity (LC₅₀ = 1.58 mg/L), followed by 3-hydroxycarbamazepine and 10,11-epoxycarbamazepine (LC₅₀ = 35.40 mg/L and 37.60 mg/L, respectively). The mixture of CBZ with its transformation products resulted in an LC₅₀ of 40.8 mg/L, suggesting a compounded toxic effect.

In addition to lethality, Oropesa et al. (2016) analyzed reproductive parameters in D. magna.Although no changes were observed in age at first reproduction, brood size, or offspring body length, significant effects were noted at the highest concentration (200 μg/L), including a 55% reduction in female reproductive output and increased abnormalities in offspring (53-61%). These malformations involved trapped eggs (linked to gastrulation failure) and curved shell spines (linked to abnormal organogenesis). Interestingly, embryos themselves did not exhibit developmental abnormalities.

Nkoom et al. (2019) contributed additional insights by evaluating bioconcentration, feeding, filtration, and neurotoxicity. At concentrations of 5 and 100 μg/L, CBZ accumulated in the organisms during the first phase of a 48h exposure period, then stabilized. Feeding and filtration rates declined with increasing CBZ concentrations, with ingestion decreasing by 11-70% and filtration by 16-78%, in a dose-dependent manner. Additionally, acetylcholinesterase (AChE) activity was significantly inhibited (53-77%), indicating a neurotoxic effect of CBZ in D. magna.

Daphnia similis

Chen et al. (2019a) investigated both acute (4-day) and chronic (21-day) CBZ exposure in Daphnia similis. During acute exposure (6.25-200 μg/L), mortality remained below 3%, and molting was not significantly affected until 200 μg/L, where it was reduced after 48-96 h. No changes were detected at 24 h.

Under chronic exposure (21 days) to different concentrations varying between 0.03 and 30 μg/L, mortality remained below 2%, but molting was significantly affected at concentrations ≥3 μg/L. The average number of molts in the control was 15.50 ± 1.38, while values for 3 and 30 μg/L were 10.17 ± 0.75 and 6.83 ± 0.75, respectively. Although the time to first brood was unchanged, brood size was significantly reduced at 30 μg/L. Furthermore, average broods per female, offspring per brood, and total offspring per female were all negatively affected by CBZ, underscoring its impact on fecundity even at low concentrations.

Corbicula fluminea

Chen et al. (2021) studied the effects of CBZ on the freshwater bivalve Corbicula fluminea, focusing on genes related to the GABAergic system (GABARAP, GABARAPL2, and GAT-1), a key component of the neuro-endocrine-immune axis. Gene expression was analyzed in multiple tissues (mantle, gill, gonad, digestive gland) after exposure to 0.5, 5, and 50 μg/L CBZ.

GABARAP expression was unchanged in the mantle and gills at 0.5 μg/L but was downregulated at higher concentrations. In the gonads and digestive glands, transcript levels decreased at all concentrations, with the lowest expression recorded in these tissues. GABARAPL2 expression was reduced in all tissues, except the gills, at the lowest concentration. In contrast, GAT-1 was upregulated in all tissues, with the highest expression in the digestive gland. These findings suggest that CBZ disrupts neuroregulatory gene expression in a tissueand dose-specific manner.

Dreissena polymorpha

Baratange et al. (2023) investigated the effects of CBZ and its co-exposure with methylmercury (MeHg) in the bivalve Dreissena polymorpha. At both time points analyzed (24 h and 72 h), co-exposure altered metabolites and proteins involved in key processes such as synaptic transmission and energy metabolism. Compared to individual exposures, the combined treatment modulated a higher number of proteins-108 in total-whereas CBZ and MeHg alone affected 46 and 55 proteins, respectively.

Regarding animal metabolite alterations, 7 and 9 compounds were affected by CBZ and co-exposure, respectively, at 24 h. At 72 h, co-exposure altered 10 metabolites. These metabolites were primarily involved in aminoacyl-tRNA biosynthesis and valine, leucine, and isoleucine metabolism and degradation. Additionally, CBZ alone induced specific changes in glycine, serine, and threonine metabolism, as well as biotin metabolism at 24 h.

Molecular and enzymatic defense responses, including lipid peroxidation in the digestive glands, gills, and gonads, were also evaluated. No significant modulation was observed in any treatment group when compared to the control, suggesting that the species' defense mechanisms were effective under the experimental conditions (CBZ: 6.1 ± 0.2 μg/L; MeHg: 460 ± 20 ng/L).

Pseudokirchneriella subcapitata

Zind et al. (2021) assessed the ecotoxicity of CBZ and its metabolites in the freshwater alga Pseudokirchneriellasubcapitata.Dibenzazepine exhibited the highest toxicity (EC₅₀ = 5.29 mg/L), followed by CBZ itself (EC₅₀ = 10.40 mg/L). 3-Hydroxycarbamazepine also showed relevant effects (EC₅₀ = 28.70 mg/L). The combined exposure to CBZ and its degradation/metabolism products resulted in an EC₅₀ of 37.0 mg/L, indicating moderate toxicity when compared to the individual compounds.

Schmidtea mediterranea

Ofoegbu et al. (2019) investigated the effects of CBZ on the flatworm Schmidtea mediterranea at concentrations of 0.01, 0.1, 1, and 10 μg/L. The drug did not induce mortality after 96 h nor did it affect feeding activity at any tested concentration. However, locomotor activity was stimulated at the intermediate concentrations (0.1 and 1 μg/L). Other parameters, such as head regeneration time, asexual reproduction, and DNA damage, remained unaffected by CBZ exposure.

Eriocheir sinensis

Chen et al. (2019b) examined the effects of CBZ on the Chinese mitten crab Eriocheir sinensis exposed to concentrations of 0.01, 0.1, 1, and 10 μg/L. Chitinase activity, a key enzyme in the molting process, was significantly reduced at 10 μg/L (to 0.55-fold relative to the control), while chitobiase was significantly inhibited at both 1 and 10 μg/L. Levels of the molting hormone 20-hydroxyecdysone (20E), an important biomarker of endocrine disruption, were also reduced at 10 μg/L (0.83-fold compared to control).

Additionally, CBZ exposure altered the expression of molting-related signaling genes. Expression of chh and mih increased at 1 and 10 μg/L (1.71-3.85 fold), while ecr and rxr were downregulated, particularly at 10 μg/L (0.22 and 0.17 fold, respectively). Chronic exposure (40 days) did not significantly affect mortality, but growth and molting were inhibited at 1 and 10 μg/L. These findings indicate that CBZ can interfere with hormonal and molecular pathways critical to development and physiological regulation in crustaceans.

DISCUSSION

The data analysis indicates that all evaluated species exhibited some level of response to exposure to carbamazepine (CBZ) and/or its metabolites. A summary of the reported effects for each species is presented in Table I. These effects may vary according to the concentration of the drug or be independent of it, depending on the species analyzed. The zebrafish (Danio rerio) stood out as the most studied freshwater species regarding the effects of CBZ. However, comparisons of studies examining the same responses reveal conflicting results within the same species, highlighting the complexity of this type of research. A notable example is the inconsistency in embryo hatching time findings-while some studies reported a reduction, others found no significant changes in this parameter.

TABLE I
Species found in the review and their respective observed effects reported

Species	Observed Effects	Concentrations	Authors
Danio rerio	Hatching alterations, gene expression modifications, behavioral changes, morpholo-gical anomalies, reproductive effects, enzymatic activity shifts, oxidative stress, immunological responses.	1-100 pg/L	Rodrigues et al. (2023); Chen et al. (2020); Zhou et al. (2019); Qiang et al. (2016); Yang et al. (2023); Jia et al. (2020); Fraz et al. (2018); Da Silva Santos et al. (2018); Weichert et al. (2017)
Cyprinus carpio	Lethality, bioaccumulation, lipid peroxidation, enzymatic activity changes.	LC50 ≈ 42.6 mg/L; 147.42 pg/g (tissues)	Gheorghe etal. (2016); Gasca- -Pérez et al. (2019)
Pimephales promelas	Toxicity assessment.	LC50: 3.27-41.50 mg/L	Zind et al. (2021)
Jenynsia multidentata	Behavioral changes and stress responses.	10-200 pg/L	Calcagno et al. (2016)
Gobiocypris rarus	Endocrine alterations and gene expression modifications.	1; 10; 100 pg/L	Yan et al. (2018)
Tinca tinea	Oxidative stress responses.	60 pg/L (isolated); 0.02-60 pg/L (mixture)	Stancova et al. (2017)
Daphnia magna	Toxicity, neurotoxicity, repro-ductive effects.	LC50~1.58-111 mg/L; sublethal tests: 5-100 pg/L; reduction at 200 pg/L	Gheorghe et al. (2016); Oropesa et al. (2016); Zind et al. (2021); Nkoom et al. (2019)
Daphnia similis	Molting frequency variations and fecundity changes.	Acute: 6.25-200 pg/L; Chronic: 0.03-30 pg/L	Chen et al. (2019a)
Corbicula fluminea	Neuroendocrine disruptions, gene expression modifications, bioaccumulation.	0.5, 5, 50 pg/L; Bioaccumula-tion: 1-10 pg/kg	Chen et al. (2021); Burket et al. (2019)
Dreissena polymorpha	Proteomic and metabolic distur-bances.	6.1 ±0.2 pg/L	Baratange et al. (2023)
Pseudokirchneriella subcapitata	Growth inhibition, algal toxicity.	EC50: 5.29-37.0 mg/L	Zind et al. (2021)
Schmidtea mediterránea	Locomotion stimulation.	0.01-10 pg/L	Ofoegbu et al. (2019)
Eriocheir sinensis	Molting process impairments, enzymatic activity shifts, hormo-nal alterations.	0.01; 0.1; 1; 10 pg/L	Chen et al. (2019b)

Among the most recurrent toxic effects observed in this review, morphological and behavioral changes stand out, recorded in more than 46% of the species exposed to CBZ. Additionally, signs of oxidative stress were reported in approximately 30% of the organisms analyzed, covering a range of trophic levels and reinforcing the potential harmful effects of carbamazepine on aquatic ecosystems. In particular, several studies involving Danio rerio indicated that the drug compromises the animals' natural antioxidant defense mechanisms, increasing their vulnerability to oxidative stress and, consequently, the risk of tissue damage (Jia et al., 2020).

In the analyzed studies, CBZ concentrations above 100 µg/L were predominantly employed in experiments with fish, while lower concentrations were applied to invertebrates and smaller organisms, evidencing their lower resistance to the drug compared to vertebrates. Differences in LC₅₀ and EC₅₀ values across studies also likely reflect both natural biological variability (e.g., species sensitivity, life stages) and methodological inconsistencies, such as exposure duration, measured endpoints, compound purity, and environmental parameters.

Although CBZ shows moderate acute toxicity in comparison to other pharmaceuticals like diclofenac or fluoxetine, it is important to note that chronic and sublethal effects-such as alterations in reproduction, neurochemical activity, gene expression, and behavior-occur at concentrations frequently found in the environment. Combined with CBZ’s high persistence and poor biodegradability, this highlights its significant ecotoxicological relevance despite relatively high LC₅₀ values.

CONCLUSIONS

Taken together, the findings of this review highlight the negative impact of improper medication disposal -particularly of carbamazepine (CBZ)- on aquatic organisms and ecosystems. CBZ is frequently detected in freshwater environments and, although its acute toxicity may be moderate compared to other pharmaceuticals, its chronic and sublethal effects (e.g., behavioral, reproductive, oxidative, and molecular alterations) occur at environmentally relevant concentrations and affect multiple trophic levels.

This environmental contamination is aggravated by the inefficiency of conventional wastewater treatment plants, which are not designed to remove pharmaceutical residues such as CBZ. As a result, drugs and their metabolites released through domestic, hospital, or industrial effluents -and even from landfills- can reach aquatic environments and bioaccumulate in non-target organisms.

To mitigate this issue, it is essential to invest in advanced wastewater treatment technologies capable of removing emerging contaminants more efficiently. In parallel, public education and engagement initiatives are crucial to promote the proper disposal of medications and reduce pharmaceutical waste at the source.

In this context, the pharmacist plays a key role in environmental protection. Beyond ensuring the rational use of medications and adherence to prescriptions, pharmacists can contribute by:

• - Leading or supporting awareness campaigns on correct pharmaceutical disposal;

• - Encouraging the population to use reverse logistics systems for returning expired or unused drugs;

• - Collaborating with policymakers in the development and implementation of public policies aimed at sustainable pharmaceutical waste management.

By actively engaging in these practices, pharmacists help prevent the release of active pharmaceutical ingredients into the environment, thereby minimizing ecotoxicological risks and protecting aquatic biodiversity.

DATA AVAILABILITY STATEMENT

Use of data not disclosed.

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Edited by

• Associate Editor:
  Anibal de Freitas Santos Jr.

Publication Dates

• Publication in this collection
  25 Sept 2026
• Date of issue
  2026

History

• Received
  17 Feb 2025
• Accepted
  12 July 2025